Though it Is known that cardioprotection enhances myocardial survival and stem cell (SC) recruitment, whether the gene/miRNA networks involved in cardioprotection promote SC-mediated cardiac repair is unknown. Moreover, whether miRNA exert their effects primarily in myocardial cells or in injected SC, and to what extent transfer of miRNA from one cell type to another is required for cardiac repair is unknown. This critical gap in knowledge prevents development of therapeutic strategies employing manipulation of protective gene/miRNA networks and SC. The central hypothesis is that expression of protective genes and repression of apoptotic genes, mediated by the gene/miRNA network, underlies enhanced myocardial survival, improved survival and function of injected mesenchymal stem cells (MSC), and positive effects upon the heart post-MI. The rationale is that by addressing this hypothesis, we will learn which miRNAs are effective, the site and mechanism of their action. We propose two SPECIFIC AIMS; 1) Determine the role and mechanism of NF-?B-dependent microRNAs that regulate pro-survival genes in cardioprotection and MSC survival/regeneration, and 2) Determine the role and mechanism of NF-?B-dependent microRNAs regulating pro cell death genes in cardioprotection and MSC survival/regeneration. The approach is gain-and loss-of-function in cardiac cells and MSC, and in vivo, using murine models of I/R injury. Labelled RNAs and real-time video imaging will allow study of miRNA transfer from one cell to another, and the results of this at the gene expression and functional levels assessed. Cardioprotection, delivery of RNA therapeutics, and SC are being pursued in clinical trials. However, the specific combinations of these studied herein, have not been explored but are feasible. The proposed research is highly significant in that it will allow us to understand the regulatory role of miRNAs and their target genes, and to employ this knowledge in identifying therapeutic targets and developing small RNA therapeutics. This proposal is innovative in that is addresses a novel hypothesis, based upon our unique preliminary data, and incorporates the novel concept that activation of protective pathways in the myocardium will enhance survival/function of injected SC. We leverage our unique knowledge of gene/miRNA networks that result in cardioprotection for the purpose of protecting MSC in the inhospitable environment of the ischemic heart. Other novel aspects are, investigation of the role played by miRNA transfer from cell-to-cell, and use of an innovative non-viral in vivo transfection technology for small RNAs.